Tackling Babesia in South African cattle with a revamped vaccine.

Tackling Babesia in South African cattle with a revamped vaccine..

Introduction. Babesia bovis and Babesia bovine is an intracellular parasite of erythrocyte of humans and cattle. Babesiosis infection rate in cattle in South Africa is between 30-40% depending on Babesia spp. Cattle can be reservoirs of the disease for several months. Life cycles uses both cattle(host) and ticks(vectors). When present inside the erythrocyte, it cannot be detected by the host’s immune system, this is its evasion mechanism, increasing the immune response time, longer immune response times, leading to worsering symptoms..

Life cycle targets. -_iiii. Recombinant vaccine based the on Apical Membrane Antigen-1 (AMA-1) of B. bovis and B. bigemina produced and tested for its effectiveness in cattle. AMA-1 is in both Babesia spp. and it is involved in the invasion process of and egression out of the erythrocyte as a merozoite. Thus targeting it will decrease anaemia symptoms, and also making the cattle less likely to remain infectious after recovery. RAP-1 is another surface protein that helps with invasion of the erythrocytes. T argeting it as well will increase the effectiveness of vaccine.

Problem statement. When ticks attach to the epidermis of the cattle, the B. bigemina and B. bovis , the tick will deposit sporozoites into the blood flow of the cattle. Once inside the sporozoites target the erythrocytes, leading to reduced appetite, high temperature, and in severe cases death. The infection of cattle with the parasite results in major economic losses($21.6 million yearly) such as a decline in the production of milk, meat, and other cattle by-products. The downstream consequences are poverty and job loss for people who work in industries dependent on cattle. The prevention methods that are currently used are not effective . Previously developed vaccines such as the attenuated ones have various disadvantages such as temperature sensitivity and difficulty in manufacturing, so a new vaccine that is more temperature stable, easier to manufacture and effective is required. In this research, we will explore the use of recombinant-based vaccines as an alternative method. These vaccines develop antibodies against the specific antigens of a disease, we are going to target the merozoite stage of the parasite infection cycle..

Aim and hypothesis. Null hypothesis: Shared antigens; AMA-1 and RAP-1 of are both present in B.bigemina and B.bovis . They a re capable of providing immunity against both types species. By stimulating release of IFN-? cytokines and TNF-? cytokines. Research hypothesis: AMA-1 and RAP-1 do not induce IFN-? and TNF-? cytokines against both B.bovis and B.b igemina . The purpose of this project is to create a recombinant vaccine against B.bovis and B.bigemina using AMA-1 and RAP-1, that will induce an immune response for the cattle..

Objectives. 1. Produce a . recombinant vaccine effective against both B.Bovis and B.Bigemina using AMA-1 and RAP-1 antigens 2. Vaccination will focus primarily on calves of the species. 3. Illustration of vaccine efficacy by testing for levels of IFN-y and TNF alpha cytokines..

Methodology. 1)DNA samples generation AMA-1 and RAP-1 gene for both B.bovis and B.bigimena will be obtained from the protein database of genbank . Primers were made and recombination into a vector was done. 2) Protein synthesis Insertion into Saccharomyces cerevisiae (yeast cells) to synthesise recombinant. The substrate and nutrients are added. Crystallisation of protein to demonstrate purity of protein and correct folding occurred..

Expected Results. E xpect results are that the vaccine stimulates both innate and adaptive immune systems which will generate IgG2a antibodies due to antigen presentation to CD4+ cells. Followed Th1 cells secretion of IgG2a and cytokines such as IFN- γ and TNF- α . By having both RAP-1 and AMA-1 recombinants in the vaccine it increases the secretion for IFN- γ . As both proteins results in an immune responding by producing of IFN- γ . The results should reflect this by increased levels of IFN- γ and TNF- α pro-inflammatory cytokines above and when looking at the results for determining immunoglobulin content in we expect an increase in IgG2 and IgG1 whose production is induced by RAP-1 and AMA-1 respectively. These antibodies should recognize and respond to Babesia spp., If a strong immune response is not developed, a booster shot will be administered to increase antibody production..

Budget. Custom crude peptides are purchased at GE script starting from $5 residue to $6 residue Elisa machine and kits costs around $2000 Western blot machinery and kits costs around $3500 for a high speed semi dry system Laboratory mice are purchased around $13 per cage Clinical research trails for vaccine are estimated at around -$1.1 million from preclinical trails until phase 3 where safety and efficiency is tested on our cattle. This includes cumulative cost of failed vaccine candidates through the trials Production of vaccine would cost a minimum of $2.8 –3.7 billion.

Timeline.